Behavioral and physiological synchrony facilitate emotional closeness in attachment relationships. The aim of this pseudorandomized cross-over study was to investigate the emotional and physiological link, designated as co-modulation, between dogs and their owners. We measured the heart rate variability (HRV) and physical activity of dogs belonging to co-operative breeds (n = 29) and their owners during resting baselines and positive interaction tasks (Stroking, Training, Sniffing, Playing) and collected survey data on owner temperament and dog–owner relationship. Although overall HRV and activity correlated between dogs and their owners across tasks, task-specific analyses showed that HRV of dogs and owners correlated during free behaving (Pre- and Post-Baseline), whereas the activity of dogs and owners correlated during predefined interaction tasks (Stroking and Playing). Dog overall HRV was the only predictive factor for owner overall HRV, while dog height, ownership duration, owner negative affectivity, and dog–owner interaction scale predicted dog overall HRV. Thus, the characteristics of dog, owner, and the relationship modified the HRV responses in dog–owner dyads. The physiology and behavior of dogs belonging to co-operative breeds and their owners were therefore co-modulated, demonstrating physiological and emotional connection comparable to those found in attachment relationships between humans.